EFFECTS OF VARIOUS VITAMIN B6 5'-PHOSPHATE DERIVATIVES ON STRUCTURE AND ACTIVITIES OF L-ASPARTATE BETA-DECARBOXYLASE

Pyridoxal 5′-phosphate and other vitamin B6 5′-phosphate derivatives (2-norpyridoxal, 2-nor-2- ethylpyridoxal, 2-nor-2-n-butylpyridoxal, 3-O-methylpyridoxal, N-methylpyridoxal, pyridoxine, pyridoxamine, and 4′-deoxypyridoxine) induce association of the low molecular weight apoenzyme (s20,w = 6 S) of laspartate β-decarboxylase to a form of the enzyme of much higher molecular weight (s20,w = 19 S). Close to 1 mole of vitamin B6 derivative is bound per minimal catalytic unit, in general with high affinity. Association does not require Schiff base formation between vitamin B6 derivative and enzyme. The N-methylpyridoxal 5′-phosphate-enzyme, shown here not to have such a Schiffbase linkage, catalyzes at low but significant rates the transamination and the β-decarboxylation of D-aspartate. The product of D-aspartate decarboxylation is L-alanine. These findings suggest that absence of a Schiffbase linkage between coenzyme and enzyme leads to a less restricted substrate binding site, and are in accord with intermediate formation of a ketimine lacking an asymmetric α-carbon atom. With other vitamin B6 derivatives, the enzyme exhibits strict L specificity. Replacement of the 2-methyl of pyridoxal by a hydrogen atom, ethyl group, or n-butyl group results in greatly decreased decarboxylase and desulfinase activities. The Km value for cysteinesulfinate is not affected by replacement of the C2-methyl by a hydrogen atom or ethyl group. The decarboxylase and desulfinase activities of the 3-O-methylpyridoxal and N-methylpyridoxal forms of the enzyme are also relatively low. In contrast, the L-aspartate-pyruvate transaminase activities observed with the 2-nor, 2-nor-2-ethyl, 2-nor-2- butyl, 3-O-methyl, and N-methyl derivatives are much higher than or 65-76% as high as that of the pyridoxal 5′-phosphate-enzyme. The substantial transaminase activity catalyzed by the N-methylpyridoxal 5′-phosphateenzyme suggests that little catalytic advantage can be ascribed to a trans-Schiffization mechanism. © 1969, American Chemical Society. All rights reserved.